Methods and Compositions for Increasing Storage-Life of Fruit

Abstract

The invention provides methods and compositions for producing plants with fruit having increased post-harvest storage life, the method comprising reducing the expression or activity in the plant, of a polypeptide with the amino acid sequence of SEQ ID NO: 1, or a variant of the polypeptide. The invention provides host cells, plant cells and plants transformed with the polynucleotides of the invention. The invention also provides methods for selecting plants with fruit having increased post-harvest storage life. The invention also provides plants produced and selected by the methods of the invention.

1 . A method for producing fruit having, or a plant with fruit having increased firmness during or after post-harvest storage the method comprising reducing the expression or activity, in the fruit or the plant, of a polypeptide with the amino acid sequence of SEQ ID NO: 1, or a variant of the polypeptide with at least 90% identity to the amino acid sequence of SEQ ID NO: 1, wherein the variant has polygalacturonase activity, wherein the method comprises the step of introducing a polynucleotide into a plant cell, the fruit or the plant to reduce the expression of the polypeptide or variant, wherein the increased firmness is relative to that of a control fruit under the same conditions, and wherein the fruit or plant is from a  Malus  species. 
     
     
         2 . The method of  claim 1 , wherein the fruit produced additionally has at least one of:
 a) reduced water loss,   b) reduced cell separation,   c) increased juiciness,   d) increased crispiness;   e) increased waxiness, and   f) reduced susceptibility to necrophytic pathogens,   during, or after, post-harvest storage, relative to that of the control fruit under the same conditions.   
     
     
         3 . The method of  claim 1 , wherein the polynucleotide comprises 21 contiguous nucleotides complementary to part of an endogenous gene, or nucleic acid, that encodes the polypeptide or variant thereof. 
     
     
         4 . The method of  claim 3 , wherein the endogenous gene comprises at least one of:
 a) a sequence with at least 90% identity to the sequence of SEQ ID NO: 4,   b) the sequence of SEQ ID NO: 4,   c) a sequence with at least 90% identity to the sequence of SEQ ID NO: 5, and   d) the sequence of SEQ ID NO: 5.   
     
     
         5 . The method of  claim 1 , wherein the polynucleotide is introduced into the plant as part of a genetic construct. 
     
     
         6 . The method of  claim 5 , wherein the genetic construct is an expression construct comprising a promoter operably linked to the polynucleotide. 
     
     
         7 . The method of  claim 6 , wherein the polynucleotide is in an antisense orientation relative to the promoter. 
     
     
         8 . The method of  claim 1 , wherein the polynucleotide comprises at least one of:
 a) at least 21 contiguous nucleotides of a sequence with at least 90% identity to the sequence of SEQ ID NO: 4,   b) at least 21 contiguous nucleotides of the sequence of SEQ ID NO: 4,   c) at least 21 contiguous nucleotides of a sequence with at least 90% identity to the sequence of SEQ ID NO: 5, and   d) at least 21 contiguous nucleotides of the sequence of SEQ ID NO: 5.   
     
     
         9 . The method of  claim 1 , wherein the plant with reduced expression of the polypeptide is regenerated from the plant cell. 
     
     
         10 . A fruit or plant produced by the method of  claim 1 , wherein the fruit or plant is genetically modified to contain the polynucleotide. 
     
     
         11 . A fruit comprising, or a plant producing fruit comprising an expression construct comprising a promoter operably linked to a polynucleotide comprising at least one of:
 i) a fragment of at least 21 contiguous nucleotides of a sequence with at least 90% identity to any one of SEQ ID NO: 4, 5, 6 and 7, wherein the sequence with 90% identity to any one of SEQ ID NO: 4, 5, 6 and 7, encodes a polypeptide with polygalacturonase activity, and wherein the polynucleotide is in an antisense orientation relative to the promoter, and   ii) a fragment of at least 21 contiguous nucleotides from any one of SEQ ID NO: 4, 5, 6 and 7,   wherein the promoter is heterologous to the polynucleotide, wherein the fruit has increased firmness during or after post-harvest storage relative to that of a control fruit under the same conditions and wherein the fruit or plant is from a  Malus  species.   
     
     
         12 . The fruit or plant of  claim 11 , wherein the expression construct is an RNAi construct. 
     
     
         13 . The fruit or plant of  claim 11  that has reduced expression of an endogenous nucleic acid corresponding to the polynucleotide. 
     
     
         14 . The fruit or plant of  claim 13 , wherein the endogenous nucleic acid encodes a polypeptide with polygalacturonase activity. 
     
     
         15 . The fruit or plant of  claim 11 , wherein the fruit additionally has at least one of:
 a) reduced water loss,   b) reduced cell separation,   c) increased juiciness,   d) increased crispiness,   e) increased waxiness, and   f) reduced susceptibility to necrophytic pathogens,   during, or after, post-harvest storage, relative to that of the control fruit under the same conditions.   
     
     
         16 . A plant part, seed, fruit, propagule or progeny of the plant of  claim 10  that is genetically modified to contain the polynucleotide. 
     
     
         17 . A plant part, seed, fruit, propagule or progeny of a plant of  claim 11  that is genetically modified to contain the construct. 
     
     
         18 . The method of  claim 1 , wherein the level of the polypeptide or variant in the fruit is less than 10% of that in the control fruit after 16 weeks post-harvest storage at 5° C. 
     
     
         19 . The method of  claim 18 , wherein the fruit have an increase in firmness of at least 40% versus control fruit after 16 weeks post-harvest storage at 5° C. 
     
     
         20 . The method of  claim 1 , wherein the fruit have an increase in firmness of at least 40% versus control fruit after 16 weeks post-harvest storage at 5° C.